Batch biosorption studies of ammonical nitrogen (NH₃-N) ions from aqueous solutions using the ubiquitous bacteria Klebsiella sp.: equilibrium, kinetic, and thermodynamic studies

Abstract

Background

The ubiquity of ammonical nitrogen (NH₃-N) in aquatic habitats is a contradictory phenomenon since it serves a crucial function in maintaining these ecosystems, yet when levels are too high, they can have adverse effects on ecological balance and human welfare. An extensive set of batch tests were used in this study to see how well the bacterial species Klebsiella sp. broke down ammonical nitrogen (NH₃-N).

Results

The research results established that Klebsiella sp. has a remarkable capacity to adapt to ammonical nitrogen concentrations of up to 125 mg/l over a long period of time. The adaptation process depends on several factors such as biomass abundance, ammonical nitrogen concentration, pH, and temperature. This study identified the optimal method for the absorption of ammonical nitrogen (NH₃-N) from a solution at a concentration of 100 parts per million (ppm), achieving an efficiency of 89 ± 1.5% mg/g under specified conditions. At a pH of 6.5, the adsorbent dosage was 0.3 g in 50 milliliters of NH₃-N at a temperature of 26 degrees C. We used an extensive range of analytical techniques, such as Scanning Electron Microscopy, Xray diffraction, Brunauer-Emmett-Teller analysis, Transmission Electron Microscopy, and Fourier-Transform Infrared Spectroscopy, to confirm the accuracy of our results. The study also showed that the biosorption process closely followed pseudo-second-order kinetics and the Langmuir model, which propose that both physical and chemical processes were involved. The thermodynamic studies also showed that this process can happen on its own and can be used in industry.

Conclusion

This study emphasizes the great ability of Klebsiella sp. to reduce NH₃-N, providing important knowledge for water quality management and aquatic ecosystem preservation.

Graphical Abstract

Introduction

The recent observed increase in the generation and management of nitrogen-rich waste can be attributed to the widespread expansion of metropolitan areas, industrialisation, commercialisation, and agricultural activities. The presence of nitrogen is crucial for the existence of every living creature since it is an integral component of proteins, nucleic acids, and other indispensable nitrogenous molecules in the biosphere (Omar et al. 2024). However, an overabundance of ammonical nitrogen (NH3-N) resulting from fertiliser residues, home waste, agricultural runoff, and animal waste can lead to toxic effects. Elevated concentrations of ammonical nitrogen can result in eutrophication, which disturbs aquatic habitats, generates unpleasant odours, and presents health hazards (Sajjad et al. 2023). Elevated concentrations of ammonical nitrogen in potable water can have significant acute and chronic health consequences for humans. Exposure to elevated concentrations of ammonical nitrogen, which includes ammonical nitrogen (NH₃) and ammonium (NH₄⁺), has substantial health hazards. Absorption of water contaminated with ammonical nitrogen can lead to gastrointestinal problems, whereas inhalation of ammonical nitrogen vapours can cause irritation to the respiratory system and eyes. Extended exposure could result in neurological after effects and breathing difficulties. Ammonium present in drinking water can elevate the risk of methemoglobinemia, often known as blue baby syndrome, in newborn babies.

Ammonical nitrogen (NH₄⁺-N) is a prevalent contaminant in aquatic ecosystems, mostly arising from agricultural runoff, commercial and industrial sewage discharge, and the breakdown of organic material. Increased concentrations of ammonical nitrogen can result in water quality degradation, eutrophication, and cytotoxicity to aquatic life, presenting a considerable risk to ecosystems and human health. The widespread distribution of ammonical nitrogen in wastewater necessitates effective treatment procedures to alleviate its detrimental impacts and also to adhere to existing environmental standards.

The release of effluents polluted with ammonical nitrogen (NH3-N) further exacerbates the degradation of water quality. Thus, it is deemed inappropriate for both human ingestion and industrial applications. Therefore, regulatory bodies have established precise criteria for water quality and acceptable levels of ammonical nitrogen exposure (Edwards et al. 2024). In the United States, acute ammonical nitrogen contamination is classified by the Environmental Protection Agency (EPA) as a concentration of 17 mg/L, while long-term intake is defined as 1.9 mg/L. Furthermore, other countries have established their own criteria for the acceptable level of ammonical nitrogen in water, ranging from 2.18 mg/L to 12 mg/L. The aforementioned requirements are applicable to pH levels ranging from 6.5 to 9.0 and encompass temperatures up to 30 °C.

Recent years have seen a notable surge in research efforts aimed at identifying effective techniques for the removal of ammonical nitrogen (NH3-N) from wastewater. To achieve this goal, traditional methods such as ammonical nitrogen stripping, chemical precipitation, and membrane separation have been widely employed. Nevertheless, they are not devoid of their disadvantages. These approaches frequently face constraints regarding the effectiveness of removal or involve significant processing expenses. Although each approach has its own advantages, they also present complexities such as efficiency issues, financial consequences, or possible environmental effects. Consequently, there is an increasing demand for creative and environmentally-friendly options to achieve more efficient removal of ammonical nitrogen from wastewater while reducing its associated disadvantages (Samy et al. 2022).

Biochemical treatment is a very effective and cost-efficient approach for eliminating ammonical nitrogen and other nitrogen-based compounds from wastewater. An ingenious approach involves immobilizing microbial biomass, hence improving the uptake of ammonical nitrogen. Furthermore, several recent investigations have investigated mordenite-chitosan and its derivatives (Safie and Yaser 2024), as presented in Table 1. In this study, Klebsiella sp. exhibited superior biosorption capacity, enhancing its efficacy in nutrient recovery. This is particularly applicable in real-world scenarios that necessitate a significant reduction of NH₄⁺. This is despite the fact that mordenite-chitosan offers advantages in terms of stability and usability, which could make it more suitable for use in large-scale industrial applications. It is possible that these advantages will increase its degree of acceptability for application in industrial activities. The combination of mordenite and chitosan may be more favourable than other materials because of its longevity and its ability to align with predictive models, which is vital for the development of new technologies for wastewater treatment. When considering the fact that mordenite-chitosan may continue to be more practicable, the comparison underlines the benefits of employing biological systems such as Klebsiella sp. for increased adsorption. When it comes to ammoniacal nitrogen biosorption, Klebsiella sp. has a number of benefits over zeolite. It efficiently eliminates ammonium ions using a dual mechanism of adsorption and metabolic assimilation, increasing overall efficiency. It has a high adsorption capacity under contrast to zeolite, which only uses physical and chemical adsorption, Klebsiella sp. continues to function consistently under a greater variety of environmental circumstances, including temperature and pH. Because it self-replicates and does not require periodic regeneration or replacement, it is also economical and renewable. Additionally, even when competing ions are present, Klebsiella sp. effectively removes ammonium ions because to its higher specificity and selectivity. Since no harsh chemicals are needed for regeneration, its environmentally benign nature lowers secondary pollutants. Furthermore, the nitrogen-enriched biomass can be recycled into biofertilizer, which is an advantage over used zeolite, which frequently needs to be disposed of. Klebsiella sp. is a better option for ammoniacal nitrogen biosorption because of these characteristics.

Biosorption is a physicochemical method that employs biological materials such as microbial biomass (bacteria, fungi, and algae) either in a viable or non-viable state to adsorb heavy metals, nutrients, dyes, and other contaminants from aqueous solutions. This process involves the passive uptake of contaminants onto the surface of biological material by mechanisms including ion-exchange, complexation, coordination, chelation, and physical adsorption. Biosorption has emerged as a feasible and environmentally sustainable alternative to conventional wastewater treatment methods for pollutant removal, especially in cases of low pollutant concentrations where traditional techniques are ineffective or too costly.

The innovation of the present study is in employing Klebsiella sp. for ammonical nitrogen biosorption, a topic that has been inadequately investigated in previous studies. This study carefully examined the biosorption capacity of Klebsiella sp. under optimum settings, including pH, the beginning ammonical nitrogen concentration, contact duration, and biomass dose, while previous research has concentrated on other microbes for nutrient removal. This objective of this study was to evaluate the biosorption capacity of Klebsiella sp. for the removal of ammonical nitrogen (NH₄⁺) from aqueous solutions, focusing on the effects of various process parameters on biosorption efficiency. The work offers novel insights into the utilization of Klebsiella sp. for efficient ammonical nitrogen removal, aiding the advancement of environmentally conscious wastewater treatment systems.

Chemical barrier serves to shield microorganisms from high levels of ammonical nitrogen and facilitates the gathering and reutilization of biomass. The ability of bacterial biomass to absorb ammonical nitrogen is influenced by several factors, including pH levels, temperature, ammonical nitrogen concentration, and the amount of available bacterial substrate (Ye et al. 2021). The primary objective of this study was to assess the capacity of Klebsiella sp., a bacterium with a rod-shaped gram-positive structure, to selectively assimilate ammonical nitrogen in various environmental circumstances. The analysis primarily investigated the influence of variables such as temperature, pH, adsorbent dosage, presence of other contaminants, and various process parameters on the adsorption capacity of Klebsiella sp. This bacterium was selected due to its validated capacity in biosorption, particularly in the removal of pollutants from wastewater. Furthermore, the system’s capacity to adjust to various ecological environmental conditions and produce extracellular polymeric substances (EPS) that serve as trapping sites for contaminants rendered it a suitable subject for our investigation.

An investigation of the correlation between dynamic forces and the uptake of ammonical nitrogen by Klebsiella sp. is of considerable significance. Eutrophication presents significant threats to aquatic environments, in which ammonical nitrogen is a prevalent pollutant. Examining the absorption capabilities of Klebsiella sp. yields pertinent insights on an ecologically sustainable approach to address eutrophication in water reservoirs. Furthermore, it is crucial to analyse the rate and efficiency of ammonical nitrogen removal in order to enhance remediation techniques. Implementing bioremediation to address ammonical nitrogen contamination not only aids in the conservation of the environment but also mitigates health hazards linked to elevated ammonical nitrogen levels, including respiratory problems, eye and skin irritations, and potential long-term effects on organ function (Li et al. 2020).

This study assessed the biosorption ability of Klebsiella sp. for the biosorption of ammonical nitrogen (NH₄⁺) from solution in water. The research entailed the isolation and characterisation of Klebsiella sp., succeeded by batch tests to investigate the influence of critical process parameters, such as pH, beginning ammonical nitrogen concentration, contact duration, and biomass dose. The results offers an in depth understanding of the biosorption efficacy of Klebsiella sp., aiding its prospective use in ammonical nitrogen eradication from wastewater (See Fig. 1).

Materials and methods

Collection, and isolation of bacteria from Waste water effluents

A series of aquatic samples were obtained from various sites and afterwards preserved at a temperature of 4 °C prior to analysis. High-quality components sourced from Sigma Aldrich were used to establish an ideal microbiological growth environment, facilitating robust microbial development. Microbiological strains were isolated from the outflow samples by inducing their growth in a culture medium supplemented with Luria-Bertani (LB) bacteria (Teshome et al. 2020). Using the traditional pour plate technique, the effluent samples were diluted and then placed onto LB agar plates. As shown in Fig. 2(a), the plates containing the bacterial culture were incubated at 37 °C for 48 h.

The post-incubation phase involved selecting several representative colonies from several culture media and analyzing their features such as morphology, biochemistry, and DNA content. The present methodological approach enabled the successful isolation and identification of bacterial strains that inhabit the wastewater samples. These strains exhibited promising potential for bioremediation applications, particularly in the context of ammonical nitrogenabsorption.

Genomic DNA purification

The genomic DNA purification protocol tracked a specific methodology as outlined (Shetty 2020). Microbial specimens were first fostered in a medium, comprised of LB (Luria broth) composition with continuous agitation at 37 °C through the night. The grown cells subsequently endured centrifugation to and the genetic material in the precipitate separated using GET solution. Hydrolytic enzymes were added and the mixture was left to incubate at 37 °C for 30 to 40 min After adding 10% SDS, the liquid extraction procedure ran at 37 °C approximately a maximum of two hours. Equivalent proportions of PCI (phenol, chloroform, and isoamyl alcohol) were used in the extraction procedure. Thirty liters of TE (Tris-EDTA buffer) were applied to disintegrate the particles that formed following the addition of ethanol to the resulting lysate. The determined genome-wide DNA was subsequently examined using 1% agarose gel chromatography.

The complex methodology enabled the effective isolation and quality control of genetic DNA from bacterial cultures derived from the wastewater samples. The visualization of DNA on an agarose gel confirmed the effectiveness of the extraction procedure and allowed for the evaluation of both the quality and amount of the isolated DNA before conducting additional analysis or identifying the bacterial strains (Maghini et al. 2021).

Amplifying 16 S rDNA via PCR

The popular primers 26 F (5′-CCAGTCCGATCMTCGCTCG-3′) and 1439R (5′GGTTACCTTGTTACGACCC-3′) were used to intensify the 16 S rRNA gene. We made copies of the 16 S rDNA from the genomic DNA we extracted. The purity of the nucleic acid copy was 100 nanograms per liter (ng/L). We used 3.5 µL of 10X PCR buffer,1.5 mM of MgCl2, a single unit of the Taq polymerase enzyme, 100 ng of each of the primers, both forward and reverse, for an overall concentration of 25 µL for our PCR product mix. Our thermocycling profiles details are provided in Table 1. When we ran agarose gel electrophoresis, we saw a clear band at 1500 base pairs from our PCR amplicon.

Subsequently, we dispatched the purified amplicons to Barcode Diagnostics Inc. (Bangalore, India) for sequencing. We aligned the forward and reverse sequences to create a consensus 16S ribosomal RNA gene. This sequence was subjected to BLAST testing toward the NCBI GenBank ‘nr’ database (Yang et al. 2020). The Clustal W multiple alignment software was used to align the top ten sequences with the greatest consistency scores. This alignment produced an intersection matrix and an evolutionary tree using MEGA 10.

The genetic information gathered from this particular isolate is now stored within the GenBank section of the NCBI database with the accession code ON358108, confirming it as Klebsiella sp. This step-by-step process involving PCR amplification, sequencing, BLAST analysis, and phylogenetic analysis led to accurately identifying the bacterial isolate wastewater as Klebsiella sp. The Genomic (DNA) extraction pictorial representation is depicted in (Fig. 1).

Fig. 1

DNA extraction sequence

Table 1 Protocols for thermal cycling in genetic research. The thermocyclic patterns for this approach involve the subsequent steps listed below

Chemical reagents

The required ingredients and laboratory-safe ammonium chloride were purchased from Sigma Aldrich (USA), supplies.

Results and discussions

Microorganism detection and separation

To extract the target bacterium, a genetic separation procedure was performed, and sequence alignment was achieved using computer methods. The Basic Local Alignment Search Tool, commonly referred to as BLAST, was developed by (Sadiq and Othman 2022). This feature allows for the quick comparison of an inquiry pattern with millions of combinations in a repository in order to find the closest match. By matching each site in the query sequence with the existing directory, positive scores are obtained, which allow for the pairing of polymorphisms and the facilitation of gap placements. However, the presence of sufficient nucleotide matches can counterbalance the negative impact of gap insertions on alignment scores. Next, these scores are turned into binary values for alignment scoring and then put into a descriptive format. The coefficient of E is an essential metric which is employed to evaluate the significance of alignments in sequence comparisons. A decrease in the associated E-value indicates a higher level of agreement between sequences in the relational database and the desired sequence, selecting the most similar repeat as the most significant outcome. (Beal et al. 2023).

Finally, a phylogenetic association diagram, shown in Fig. 2(d), was constructed using MEGA version 5, using 16 S rRNA genotypes from bacteria as the basis. The present study allowed the graphical representation of evolutionary connections and places the isolated bacterial strain within the context of closely related species. By combining DNA sequencing, BLAST analysis for sequence similarity searches, and phylogenetic tree construction, scientists effectively defined the bacterial strain and can obtain valuable information about its evolutionary relationships with other bacteria. These bioinformatics techniques are crucial in the molecular analysis and taxonomic categorization of microorganisms (Bi et al. 2024).

Analysis and characterization

The experiment began by conducting biosorption trials under standard conditions. A Spectrophotometer (PG T80 model), calibrated with a 100-ppm ammonical nitrogen stock solution, was used to quantify the levels of ammonical nitrogen in the solutions both pre- and post-adsorption. These amounts had been detected by diluting the leftovers using distilled water.

The physical attributes of this strain were examined thoroughly by conducted imaging using scanning electron microscopy (FE-SEM, Model ZEISS GEMINI SEM 500”), presented in Fig. 2(b). By using the HR-TEM (Fig. 2c) the internal biological architecture of the Klebsiella sp. membrane could be detected before and after ammonical nitrogen adsorption. Furthermore, this strain was analysed by compound microscopy to visualise the external morphology). (Fig. 2d) In addition to this FTIR characterisation was done for material identification. (FTIR, SHIMADZU.CORP00748, as depicted in Fig. 2(f).

In addition, the BET evaluation demonstrated an exterior area of 2.3560 m²/g, a typical-sized pore width of 29.06 Å, and a pore capacity of 0.0080 cm³/g for the biosorbent, demonstrating its porous nature and effective interaction with adsorbates (Fig. 2e). This in-depth characterization through advanced analytical techniques provided valuable insights into the structural, morphological, and surface properties of the Klebsiella sp. biosorbent, as well as any alterations caused by ammonical nitrogen adsorption. An analysis of this nature is crucial for the optimization of the biosorption process and the elucidation of the fundamental mechanisms at play (Pugazhendhi et al. 2018).

Fig. 2

(a) Culture forming bacteria. Figure 2(b), displays the SEM image of the Klebsiella sp. strain. Figure 2(c) The TEM image of the strain is depicted. Figure 2(d). Compound microscopy characterisation of the strain Fig. 2(e) BET Analysis and Fig. 2(f) The FTIR analysis

Influence of pH as well as temperature

The analysis involved triple observations utilising standard errors of measurement ranging from 2 to 5% of the mean, thus ensuring robust and reliable results. The impact of temperature was examined within the range of 5–40 °C, while the pH was adjusted between 1 and 10 while simultaneously maintaining other factors constant. pH was tuned using hydrochloric acid or sodium hydroxide (Papciak et al. 2024). The decrease in the ammonical nitrogen concentration displayed a continuous increase, indicating a constant biological absorption capacity that is responsive to variations in temperature, as depicted in Fig. 3(a). At temperatures above 35 °C, Klebsiella sp. showed reduced effectiveness in ammonical nitrogen breakdown, possibly owing to enzyme inactivation. With respect to the pH effects seen in Fig. 3(b), the adsorption rate reached its highest point between 6.5 and 7.8. As a pH above this threshold, the intensity steadily declined. These findings are consistent with prior research on the ideal pH range for nitrifying bacteria such as Nitrosomonas (7.8–8.6). The results emphasise the significant impact of temperature and pH on the effectiveness of biosorption and enhance our knowledge of these parameters for improving the efficient removal of ammonical nitrogen and nitrogen using Klebsiella sp. and other biosorbents. Overall, these findings offer important insights into the interaction between ambient variables and biosorption efficacy, which can motivate further research and application in wastewater treatment and ecological remediation (Mir and Rather 2024).

Influence of ammonical nitrogen dosage

The concentration of ammonical nitrogen was measured within the range of 10–100 mg/L, while maintaining a constant temperature of 30 °C and keeping all other variables unchanged. The biosorption rate in (Fig. 3c) positively correlated with the initial ammonical nitrogen concentration, reaching its maximum at around 87 mg/L. However, as the initial ammonical nitrogen concentration exceeded 89 mg/L, the rate of ammonical nitrogen removal declined significantly with each subsequent rise. The decrease observed at greater concentrations may be attributed to the suppressive effect of elevated amounts of ammonical dosage on the biological absorption process (Huang et al. 2023).

Conversely, when the concentrations of ammonical nitrogen are low, scientists hypothesise that biosorption decelerates due to challenges in substrate mobility. This phenomenon occurs due to insufficient availability of ammonical nitrogen for the biomass to bind to. The process of mobility is considered significant in the absorption of heavy metals at low concentrations, and a similar mechanism could regulate the absorption of ammonical nitrogen. The findings demonstrate that Klebsiella sp. exhibits an optimal range of initial ammonical nitrogen concentrations for effective biosorption. The presence of excessive or insufficient amounts of ammonical nitrogen initially can disrupt the rate of removal, perhaps due to factors that impede their effectiveness or difficulties in relocating substances. Accurately determining the initial level of pollution is crucial for optimizing biosorption and minimizing the presence of contaminants in various wastewater systems (Ishaq et al. 2023).

Additional wastewater contaminants can reduce bacterial activity and biosorption. Sulphates and heavy metal ions such as iron, aluminium, and zinc have been found in normal refinery effluent by chemical analysis. The ammonical nitrogen biosorption of contaminants has been studied at various doses. The experimental results showed that these pollutants had no effect on ammonical nitrogen biosorption Fig. 3(f). These toxins were chosen to be within or significantly above wastewater levels. Iron appears to have a moderately negative impact on the biosorption rate, although microorganisms exposed to a combination of such metallic ions remain unaffected. The effects of sulphates on biosorption were examined using sodium sulphate at 100–1000 mg/l. The experimental results showed that sodium sulphate at 1000 mg/l did not affect biosorption. After each experiment, heavy metal and sulphate concentrations did not vary, save for a 10% decrease in iron.

Regeneration studies

The potential reusability of the bacteria was evaluated by conducting five consecutive adsorption-desorption cycles using the same samples, as depicted in Fig. 3(d). Following the adsorption process, we isolated the bacteria by means of centrifugation. Subsequently, the isolated bacteria were subjected to treatment with various chemical solutions in order to remove the absorbed ammonical nitrogen ions. In particular, 0.1 g samples were subjected to 25 ml solutions containing either deionized water or 0.05 N HNO3. The solutions were agitated for 15 min at a speed of 25,000 rpm, after which the concentrations of the solutions were determined.

Two techniques were employed to restore the bacteria to their original condition. First, through treatment with distilled water, and second, by treatment with a 2% sodium hydroxide solution to quickly remove reactive hydrogen ions before reaching a pH of 5. Following the regeneration process, the bacteria were meticulously washed with deionized water until they reached a pH level of neutrality. Ultimately, we desiccated the regenerated bacteria at a temperature of 40 °C in order to ready them for reutilization in novel adsorption procedures. An evaluation of adsorption-desorption cycles is crucial for determining the possible reusability and cost-effectiveness of the Klebsiella sp. biosorbent in real-world applications. This assessment attempted to demonstrate that the biosorbent may be regenerated and reused several times without experiencing substantial performance degradation.

Zeta potential studies

The pHpzc is the specific charge at which the opposing charges on the material exhibit an equilibrium, resulting in an overall charge of zero. Deciphering the pHpzc is essential for comprehending the sensitivity of the surface to changes in pH, its composition, and its ability to adsorb ions with different charges. For this work, the pHpzc value was employed to establish the point of zero charge for the Klebsiella sp. adsorbent. According to the data presented in Fig. 3(e), the pHpzc level of the Klebsiella strain adsorption material is 4.4. This particular pH value implies that the whole surface of the material does not carry any extra charges. At pH values below 4.4, the surface would exhibit a positive charge, whereas at pH values above 4.4, it acquires a negative charge. At a pH of 5, which is higher than the critical pH of 4.4, the surface of Klebsiella sp. becomes negatively charged, promoting the adsorption of positively charged ions. This was advantageous for the experimental investigation on ammonical nitrogen biosorption. Quantifying the pHpzc yields vital information about the surface charge characteristics of the biosorbent and helps in adjusting pH settings to enhance the effective adsorption of the desired ions or molecules (Gu and Lan 2023).

Fig. 3

Graphs of the various factors that affect biosorption rates. (a) First and foremost, it displays how temperature plays a role. (b) Additionally, it delves into the impact of pH levels. (c) It also scrutinizes the influence of ammonical nitrogen. (d) Recovery analyses (e). Electrophoretic mobility investigations. (f). Effects of other co-existing ions on biosorption of ammonical nitrogen

Design of experiments (DOE) modelling

RSM modelling with a central composite design approach was further studied and both the two-dimensional contour graphical representations and a three-dimensional surface representation (Fig. 4a, b,c) clearly demonstrate the intricate relationship and complex interaction between temperature, ammonical nitrogen concentration, pH, and bacterial dosage in relation to the efficacy of ammonical nitrogen detoxification. Elevating the bacterial dosage to 1.2 g significantly improves the rates of ammonical nitrogen elimination, in line with previous research that has shown the beneficial effect of bacterial bead volume/concentration. The denitrification potential of the strain of interest can be enhanced by exposure to increasing amounts of ammonical nitrogen, which leads to adaptation and promotes the optimization of capacity biosorption. Initial ammonical nitrogen concentrations of moderate levels improve removal rates, loadings that are too high hinder efficiency, emphasizing the complex connection between concentration and process efficiency (Aminu et al. 2023).

The highest levels of bacterial activity occur within intermediate temperature ranges (25–37 °C), whereas significantly reduced performance is observed at extreme temperatures, which correspond to the optimal circumstances for bacterial growth. The ideal pH for the elimination of ammonical nitrogen is around 7, with progressive reductions beyond pH 7–9, and biosorption is impeded in acidic environments, thus underscoring the importance of pH optimization.

Studies on kinetics and biosorption

The present work was designed to examine the adsorption characteristics and rate of Klebsiella sp. by the utilization of rate equations that were derived from established pseudo-first-order and pseudo-second-order adsorption rates (Rahim and Adina, n.d.). The involvement of Klebsiella sp. plays a vital role in the process of ammonical nitrogen biosorption, progressing through two principal phases. Initially, the bio sequestration process begins quickly but then gradually decelerates in each following phase, resulting in a continuous increase in the rates of biosorption.(Mir and Rather 2024).

Table 2. Values of Q_e, k₁, k₂, and the coefficient of correlation (R²) obtained using graphical statistical analysis. Comparative analysis reveals that the pseudo-second-order model (R² = 0.99) has a more substantial correlation coefficient value, so providing a superior fit compared to the pseudo-first-order model (R² = 0.95). These findings indicate that the sorption of ammonical nitrogen onto Klebsiella sp. primarily occurs via biosorption mechanisms.

Table 2 Rate parameters of nitrogen-ammonical nitrogen biosorption

Subsequently, the data related to the adsorption were evaluated using two kinetic models, namely pseudo-first-order and pseudo-second-order kinetics. The models are mathematically defined by certain equations:

$$\begin{gathered} {\text{Pseudo-first-order}}\,{\text{ kinetics}}\,{\text{ model:}}\, \hfill \\ \;\;\;\;\;\;\;\;Log\left({{q_e} - {q_t}} \right)=log{q_e} - \frac{{{K_1}t}}{{2.303}} \hfill \\ \end{gathered}$$

(1)

$$\begin{gathered} {\text{Pseudo-second-order}}\,{\text{ kinetics }}\,{\text{model: }} \hfill \\ \;\;\;\;\;\;\;\;\;\;\;\;\;\;\;\;\;\;\;\;{\raise0.7ex\hbox{$t$} \!\mathord{\left/ {\vphantom {t {qt}}}\right.\kern-0pt}\!\lower0.7ex\hbox{${qt}$}}={\raise0.7ex\hbox{$1$} \!\mathord{\left/ {\vphantom {1 {\left({{K_2}{q_e}2} \right)}}}\right.\kern-0pt}\!\lower0.7ex\hbox{${\left({{K_2}{q_e}2} \right)}$}}+{\raise0.7ex\hbox{$t$} \!\mathord{\left/ {\vphantom {t {{q_e}}}}\right.\kern-0pt}\!\lower0.7ex\hbox{${{q_e}}$}} \hfill \\ \end{gathered}$$

(2)

In this paper, Q_e represents the adsorption capability, while Q_t denotes the binding capacity at a certain moment in time (t in minutes). The adsorption kinetic parameters are denoted by k₁ (min^− 1) and k₂ (in grammes of weight per milligramme per minute). The relevant dynamics are represented by the linearly adapted curve shown in Fig. 5(a) and 5(b). Based on the analysis of the correlation coefficients between these two kinetics variables, it is evident that adsorption significantly affects the rate of adsorption material absorption.

Biosorption equilibrium models can be utilized to evaluate the potential of biomass materials because they provide insights into the surface attributes and adhesion properties of the biological material. In the present study, four widely used equilibrium paradigms (Langmuir, Freundlich, Temkin, and sips isotherms) were employed to analyse the process of biosorption, as illustrated in Fig. 5(c), (d), (e), (f), respectively. The fundamental assumption behind the Langmuir isotherm model is that the absorbency mechanism operates at discrete, homogeneous spots on the absorbing material surface. It is also possible to express this model using non-linear equations, as reported in a previous study (Mushtaq et al. 2023).

$$qe=\frac{{{q_{m{K_l}}}{C_e}}}{{1+{K_l}~{c_e}}}$$

(3)

In this context, the variables “qm” and “Ce” represent the monolayer sorption efficiency (mg/g) of the material, “K_L” and the Langmuir adsorption constant (mg/L) which are associated with the free energy of sorption, and “q_e” which denotes the equilibrium metal ion concentration (mg/g) of the resin.

The Freundlich model predicts that the sorption region is heterogeneous. This diversity is predicted by Freundlich’s model (Mir and Rather 2023).

$${q_e}={K_f}{C_e}1/n$$

(4)

In this predictive models, ‘K_f’ functions as a constant representing sequestration capacity. On the other hand, ‘1/n’ serves as an attribute that expresses the level of biological absorption variation based on material characteristics. The values of ‘K_f’ and ‘1/n’ were obtained through nonlinear regression analysis. Non-linear Freundlich isotherm graphs are depicted in Fig. 5(d). readings of “1/n” that lie between 0 and 1 denote successful biosorption of ammonical nitrogen by the biomass under the specific conditions outlined in Table 3.

Table 3 Isotherm model parameters

The Temkin isotherm paradigm examines the impact of temperature variations on both the adsorbent (biomass) and the adsorbate (ammonical nitrogen). Empirical evidence indicates that elevated temperatures result in enhanced heat absorption during the adsorption process. These findings suggest that chemisorption and ionic exchange technology are more favourable approaches for elimination compared to physical absorption (Mir and Rather 2023). Previous studies have demonstrated a significant correlation between the Langmuir isotherm and a prominent ion transfer mechanism. The subsequent complex formation events that occur after adsorption are elucidated by the Freundlich isotherm. The experimental data exhibit a strong correlation with both the Langmuir and Freundlich isotherms, indicating that ion exchange and complexation play crucial roles in promoting the binding of ammonical nitrogen to Klebsiella sp. bacteria (Mir and Rather 2023).

Fig. 4

RSM. (a) Expansion plots displaying the perturbation of all four variables, (b) plot showing studentized residuals against run number. (c) Graphs of experimental values versus the anticipated values and RSM 3D contour graphs (d, e, f) and their relationship with the factors being considered

Fig. 5

Pseudo-first-order kinetics are observed in Fig. 5(a), indicating the presence of concentration-dependent interfacial attraction. Figure 5(b) illustrates chemisorption using pseudo-second-order kinetics, where the absorption rate follows an exact correlation with the square of the dosage. Figure 5(c). The Langmuir fitting method is a nonlinear technique used to determine adsorption capacity. Figure 5(d) presents an explanation of surface multilayer adsorption based on the Freundlich hypothesis. An approximate linear decrease in heat is observed at adsorbate-adsorbent interactions, as shown by the Temkin model in Fig. 5(e). A further elucidation of the Sips model is provided in Fig. 5(f). The thermodynamic analysis is shown in Fig. 5(g), which investigates changes in enthalpy, entropy, and Gibbs free energy

Adsorption mechanism

The applications of bioremediation leverages the inherent binding characteristics present in various biological materials. The approach described enhances the capacity of biological particles to absorb nutrient pollutants/heavy metals from wastewater by incorporating both metabolically mediated and physico-chemical absorption mechanisms. Due to the intricate interplay of many factors that affect certain biosorbents, it is challenging to provide a general approach. Despite the consideration of several suggested metal-binding mechanisms such as chemisorption by ion exchange, coordination and/or chelation, complexation, physical adsorption, and others, the precise mechanism of metal biosorption remains incompletely established. Nutrient pollution mostly influences the intake of nutrients through active transport facilitated by the activation of specific ionic channel proteins on their surfaces. This upregulation enables molecular trafficking and ultimately the incorporation of these ions into the cytoplasm. The hypothetical mechanism is depicted in Fig. 6.

Fig. 6

Possible mechanism of bioremediation of ammonical nitrogen in species of bacteria

Conclusions

The present work provides a thorough investigation of the exceptional effectiveness of Klebsiella sp., a bacterium, in the mitigation of ammonical nitrogen pollution in wastewater. Under specific conditions, such as a pH of 6.5, utilising 0.5 g of adsorbent biomass in a 50-millilitre ammonical nitrogen solution at a temperature of 26 degrees C, the results indicate that it has the potential to remove up to 89.1 ± 1.5% of ammonical nitrogen. Employing advanced analytical methods, we conducted a comprehensive study of the intricate mechanisms involved in this absorption process and confirmed the precision of the Langmuir, Temkin, Freundlich, and Sips isotherm models in characterising the process. In addition, by the implementation of response surface methodology, we enhanced the process parameters to significantly enhance system efficiency.

The findings of our investigation offer verification for the ability of Klebsiella sp. to function as an ecologically harmless and cost-effective biosorbent for the remediation of ammonical nitrogen contamination in wastewater. This innovative approach has immense potential for widespread application in water treatment facilities, therefore assisting in the mitigation of eutrophication and the preservation of aquatic ecosystems. The data obtained in this study not only validate the exceptional abilities of the Klebsiella sp. strain but also provide crucial insights for tackling the pressing issue of ammonical nitrogen pollution in aquatic environments. Therefore, our investigation validates Klebsiella sp. as a highly promising candidate for sophisticated wastewater treatment applications.

In order to ensure the economic viability and practical implementation of the biosorption process, future efforts should prioritise its industrialisation. In addition, the sustainability and cost-effectiveness of the procedure will be significantly improved by the implementation of effective regeneration methods for the adsorbent biomass. Moreover, the investigation of the potential synergies between this biosorption technique and other conventional wastewater treatment methods should facilitate the development of customised solutions that address site-specific challenges. It is imperative to conduct thorough environmental impact studies and life cycle evaluations to guarantee the effectiveness of the biosorption process and thus enduring its safety and sustainability by utilising Klebsiella sp. Simultaneously, it is imperative to prioritise the understanding of the fundamental genetic and molecular mechanisms that endow this strain with an exceptional biosorption capacity. Consequently, this will accelerate the development of genetic engineering techniques that are specifically designed to improve its performance.

The work described herein undoubtedly represents significant progress towards ecologically advantageous and sustainable approaches for decontaminating wastewater, with the goal of addressing the pressing global problem of ammonical nitrogen pollution. Ongoing research and advancement of the Klebsiella sp. strain has the capacity to profoundly revolutionize the wastewater treatment sector, therefore fostering a more environmentally friendly and sustainable future for our aquatic ecosystems (See Table 4).

Table 4 Comparison table